Jeremy R. Everett

Spin-echo 1H n.m.r. spectroscopy: a new method for studying penicillin metabolism

High-field, spin-echo 1H n.m.r. spectroscopy has been used to detect the diketopiperazine, (1), a new metabolite of ampicillin, (2), in rat urine.

Parameters for the Calculation of Proton NMR Chemical Shifts of Oligoribonucleotides

A set of empirical parameters which allows the prediction of the proton NMR chemical shifts at 70 C of non-exchangeable heterobase and anomeric protons in oligoribonucleotides has been constructed. The set is based on the highly flexible nature of oligoribonucleotide single strands and the wide range of conformational states which can be populated at relatively high temperatures (70 C or greater). A pairwise subtractive procedure, using 129 ribonucleotide oligomers (all 16 dimers, all 64 trimers, 37 tetramers, and 12 pentamers), shows that significant contributions to the observed chemical shift of protons in a given nucleoside residue are made by first, second, and third neighbors on the 3 and the 5 sides. The majority of the neighbors cause shielding effects with the exception of some first neighbors on the 5 side of a given residue. The magnitude of the shielding effects is greatest for the purine heterobases and follows the order A greater than G greater than C greater than U, with first neighbors on the 3side showing more pronounced effects than second neighbors and these in turn showing larger effects than third neighbors. Second neighbors on the 5 side showed consistently greater shieldings than first neighbors, a result attributed to the deshielding effects of the first 5 neighbor phosphate group. The parameter Tables are applied to the prediction of proton chemical shifts in one heptamer, four hexamers, and two pentamers and give average absolute differences between predicted and observed shifts less than 0.030 ppm. The parameter approach represents an excellent method of generating initial assignments of proton chemical shifts for any single strand oligoribonucleotide.

The conformational analysis of erythromycin A

1 n H and 13C n.m.r. spectroscopy combined with molecular modelling techniques were used to show that the major conformation of erythromycin A (1) in CDCl3 solution is very similar to the crystalline state conformation (A) of erythromycin A hydroiodide dihydrate. However, 1H nuclear Overhauser enhancement (n.O.e.), variable-temperature and variable-solvent n.m.r. experiments showed that, in contrast to previous reports, the major conformation of (1) is in fast equilibrium with a second, minor conformation. This minor conformation is related to the major conformation via a reorganisation of the macrocyclic lactone ring in the C-2 to C-9 region. n 13 n C N.m.r. relaxation measurements showed that the desosamine sugar possesses more conformational freedom than the cladinose sugar, in agreement both with previous results and with energy calculations based on the crystal structure. In addition, the relaxation experiments indicated that some methyl groups in (1) are sterically hindered whereas others possess motional freedom. A good agreement was found between the n.m.r. relaxation results and calculations of methyl group rotational energy barriers in the crystal structure. This good agreement provided further evidence of the similarity between the major solution-state conformation of (1) and the crystalline-state conformation A.

The conformational analysis of three derivatives of erythromycin A: (9S)-9-hydroxy-9-deoxoerythromycin A, (9S)-9,11-O-isopropylidene-9-deoxoerythromycin A, and (9S)-erythromycylamine A by nuclear magnetic resonance spectroscopy and molecular modelling

The 1H and 13C n.m.r. spectra of (9S)-9-hydroxy-9-deoxoerythromycin A (2), (9S)-9, 11-O-isopropylidene-9-deoxoerythromycin A (3), and (9S)-erythromycylamine A (4) were solved by two-dimensional (2D) n.m.r. methods. The solution conformations of compounds (2), (3), and (4) were then determined by n.m.r. spectroscopy [n.O.e.(1D and 2D) and T1 experiments] and compared with the solution conformation of erythromycin A (1). In each case the cladinose and desosamine sugars are in the same chair conformations as found in (1). Furthermore the overall orientations of both sugars were found to be very similar to those of (1) in all cases. However, the lactone rings of (2) and (4) were found to be in fast exchange between conformations characterised as ‘folded-in’ or ‘folded-out’ in the C-3 to C-5 region, whereas the lactone ring of (3) was predominantly in a ‘folded-out’ conformation, as was found for (1). The ‘folded-in’ conformations are novel and have not been previously described. Molecular modelling was used in order to compare the solution-state conformations with crystalline-state models of the two types of lactone ring conformation. No simple correlation was found between the solution-state conformational preferences of compounds (1)–(4) and their antibacterial activities.

The conformational analysis of derivatives of erythromycin A: X-ray crystallographic and nuclear magnetic resonance spectroscopic studies of (E)-11-O-(2-dimethylaminoethoxy)methyl-9-deoxo-9-methoxyiminoerythromycin A

(E)-11-O-(2-Dimethylaminoethoxy)methyl-9-deoxo-9-methoxyiminoerythromycin A (3) has been synthesised from erythromycin A (1) and its crystal structure conformation compared with that of (1). The conformations of the two sugars and their orientation with respect to one another are the same in (1) and (3). The conformations of the aglycones are, however, markedly different. In comparison with (1), the main conformational changes in (3) appear to be a folding in the aglyconie about the C-7 methylene and lactone ring oxygen, which moves the C-6 hydroxy and lactone carbonyl away from the substituents at C-9 and C-11, and an inward folding (towards C-11) of the C-3 to C-5 region and the attached sugars.The conformation of (3) in CDCl3 was investigated by measuring 1H vicinal coupling constants, 1H n.O.e.s, and 13C spin–lattice relaxation times. These studies show that in solution. (3) exists in a state of fast exchange between conformations of the C-3 to C-5 ‘folded out’ and C-3 to C-5 ‘folded in’ types. One of the simplest representations of this would be a rapid equilibrium between conformations similar to those found in the crystal structures of (1) and (3).

Novel long-range 1H and 13C n.m.r. isotope effects transmitted via hydrogen bonds in a macrolide antibiotic: bafilomycin A1

Novel, long-range (up to 12 bonds) isotope effects transmitted via hydrogen bonds were observed in the 1H and 13C n.m.r. spectra of bafilomycin A1(1) upon partial deuteriation of the exchangeable protons.

Novel conformations of erythromycin antibiotics: the conformational analysis of erythromycin A and (9S)-9-hydroxy-9-deoxoerythromycin A by n.m.r. spectroscopy

1 n H and 13C N.m.r. spectroscopy combined with molecular modelling has been used to characterise novel ‘folded-in’ lactone ring conformations of erythromycin A (1) and its (9S)-9-deoxo analogue (2) in CDCl3 solution.

Chemical shift-correlated two-dimensional spin echo n.m.r. Spectroscopy

An alternate two-dimensional n.m.r. pulse sequence gives the chemical shifts of directly coupled nuclei.